Colicin A

Colicin A is a type of Colicin, a bacteriocin made by E. coli which acts against other nearby E. coli to kill them by forming a pore in the membrane, leading to depolarisation of the membrane which kills the cell.

Synthesis
Colicin A is synthesised through a single operon, also containing the colicin A lysis protein encoding gene, Cal, and the ColA Colicin Immunity Protein, Colicin_Immunity_Protein. The Cal gene is also involved in the expression of the colicin A operon - without the cal gene the amount of colicin A produced decreases. The cal gene product is likely to be an activator of colicin A expression, including its own expression. The N terminal region of the cal gene product is particularly involved in the regulation of col A synthesis. A separate, unidentified, product (which may be a heat-shock protein) could also be involved, and complement the cal gene product. The expression of the col A operon provokes a shut-off of chromosomal protein expression, which is due to the expression of the lysis gene. This shut-off gives the col A production a priority over other cellular proteins in their synthesis.

Synthesis of col A starts immediately after induction. At lower temperatures, like 30oc, there is a lag phase to this synthesis, which does not end in null cal mutants. At 37-42oc the lag is very short, and a large amount of col A is produced very rapidly. The signal to speed up synthesis from the lag phase involves the cal gene product, alongside the other unidentified gene product at normal and high temperatures.

Cal has a self-regulatory role in the expression of both itself and the col A gene, which may be common to all colicin lysis proteins.

Release
Extracellular release of col A is non-specific. No mutations in the central or N terminal regions of col A were found to have any effect on the release, or on the efficiency of the release. When the three domains of col A were separated they also still continued to be released at normal levels. A mutation was also inserted in the C terminus to promote aggregation of the protein in the cytoplasm, and this also had no effect on the secretion of col A, so there is no interaction between N and C termini in the release of Col A. The process of release is therefore non-specific with respect to the colicin itself, and is dependent only on the expression of the cal gene. This causes release of col A through the non-specific permeabilisation of the cell envelope.

Uptake
Colicin A binds to the BtuB Vitamin B12 outer membrane receptor of the target cell, and uses the Tol system to translocate across the membrane, specifically TolQRAB, alongside the OmpF protein. Its use of the Tol system means that Colicin A is in the A group of colicins.

This structure shows the TolB box of colA bound to TolB.

Cytotoxic activity
Colicin A exhibits Pore Formation activity, which means that its cytotoxic domain inserts into the membrane of the target cell, resulting in the depolarisation of the cell membrane. E. coli uses the polarisation of its cell membrane to generate energy, so with this not functioning a number of energy-requiring cellular functions are inhibited, and the cell ultimately dies, after an arrest of motility within 3 minutes. ColA denatured first in urea kills the cell more quickly than natively folded ColA.

Col A when present in an E. coli cell is able to affect macromolecular synthesis throughout the cell, affecting many of the processes in the cell. One such affected system is nucleic acid synthesis; it is halted very soon after Col A is added to the system. Colicin A also has a negative impact on the availability of ATP-dependent active transports, such as some permease activities, such as the uptake of labelled isoleucine.